Development of a Self-Cloning System for Actinomadura verrucosospora and Identification of Polyketide Synthase Genes Essential for Production of the Angucyclic Antibiotic Pradimicin

1999 ◽  
Vol 65 (6) ◽  
pp. 2703-2709 ◽  
Author(s):  
Tohru Dairi ◽  
Yoshimitsu Hamano ◽  
Tamotsu Furumai ◽  
Toshikazu Oki
Keyword(s):  
Nucleotide Sequence ◽  
Polyketide Synthase ◽  
Selectable Marker ◽  
Plasmid Vector ◽  
Open Reading Frame ◽  
Antibiotic Biosynthesis ◽  
Nucleotide Sequence Analysis ◽  
Reading Frame ◽  
Cloning System ◽  
Open Reading Frame 1

ABSTRACT A self-cloning system for Actinomadura verrucosospora, a producer of the angucyclic antibiotic pradimicin A (PRM A), has been developed. The system is based on reproducible and reliable protoplasting and regeneration conditions for A. verrucosospora and a novel plasmid vector that consists of a replicon from a newly found Actinomadura plasmid and a selectable marker cloned from the Actinomadurastrain. The system has an efficiency of more than 105CFU/microgram of DNA. Using this system, we have cloned and identified the polyketide synthase (PKS) genes essential for PRM A biosynthesis from A. verrucosospora. Nucleotide sequence analysis of the 3.5-kb SalI-SphI fragment showed that ketosynthase subunits (open reading frame 1 [ORF1] and ORF2) of the essential PKS genes have strong similarities (59 to 89%) to those for angucyclic antibiotic biosynthesis.

scholarly journals Characteristics of a New Enantioselective Thermostable Dipeptidase from Brevibacillus borstelensis BCS-1 and Its Application to Synthesis of a d-Amino-Acid-Containing Dipeptide

2004 ◽  
Vol 70 (3) ◽  
pp. 1570-1575 ◽  
Author(s):  
Dae Heoun Baek ◽  
Jae Jun Song ◽  
Seok-Joon Kwon ◽  
Chung Park ◽  
Chang-Min Jung ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Nucleotide Sequence ◽  
Amino Acid Sequence ◽  
Open Reading Frame ◽  
Nucleotide Sequence Analysis ◽  
Reading Frame ◽  
E Coli ◽  
Specificity Constant ◽  
Optimal Ph

ABSTRACT A new thermostable dipeptidase gene was cloned from the thermophile Brevibacillus borstelensis BCS-1 by genetic complementation of the d-Glu auxotroph Escherichia coli WM335 on a plate containing d-Ala-d-Glu. Nucleotide sequence analysis revealed that the gene included an open reading frame coding for a 307-amino-acid sequence with an M r of 35,000. The deduced amino acid sequence of the dipeptidase exhibited 52% similarity with the dipeptidase from Listeria monocytogenes. The enzyme was purified to homogeneity from recombinant E. coli WM335 harboring the dipeptidase gene from B. borstelensis BCS-1. Investigation of the enantioselectivity (E) to the P1 and P1′ site of Ala-Ala revealed that the ratio of the specificity constant (k cat /Km ) for l-enantioselectivity to the P1 site of Ala-Ala was 23.4 � 2.2 [E = (k cat /Km ) l,d /(k cat /Km ) d,d ], while the d-enantioselectivity to the P1′ site of Ala-Ala was 16.4 � 0.5 [E = (k cat /Km ) l,d /(k cat /Km ) l,l ] at 55�C. The enzyme was stable up to 55�C, and the optimal pH and temperature were 8.5 and 65�C, respectively. The enzyme was able to hydrolyze l-Asp-d-Ala, l-Asp-d-AlaOMe, Z-d-Ala-d-AlaOBzl, and Z-l-Asp-d-AlaOBzl, yet it could not hydrolyze d-Ala-l-Asp, d-Ala-l-Ala, d-AlaNH2, and l-AlaNH2. The enzyme also exhibited β-lactamase activity similar to that of a human renal dipeptidase. The dipeptidase successfully synthesized the precursor of the dipeptide sweetener Z-l-Asp-d-AlaOBzl.

Nucleotide sequence analysis of a zein genomic clone with a short open reading frame

Gene ◽  
1984 ◽  
Vol 28 (1) ◽  
pp. 113-118 ◽  
Author(s):  
Jean C. Kridl ◽  
Jeffrey Vieira ◽  
Irwin Rubenstein ◽  
Joachim Messing
Keyword(s):  
Sequence Analysis ◽  
Nucleotide Sequence ◽  
Genomic Clone ◽  
Open Reading Frame ◽  
Nucleotide Sequence Analysis ◽  
Reading Frame ◽  
Short Open Reading Frame

scholarly journals Nucleotide sequence analysis and overexpression of the gene encoding a type III chloramphenicol acetyltransferase

1988 ◽  
Vol 252 (1) ◽  
pp. 173-179 ◽  
Author(s):  
I A Murray ◽  
A R Hawkins ◽  
J W Keyte ◽  
W V Shaw
Keyword(s):  
Sequence Analysis ◽  
Nucleotide Sequence ◽  
Cyclic Amp ◽  
Chloramphenicol Acetyltransferase ◽  
Open Reading Frame ◽  
Nucleotide Sequence Analysis ◽  
Calculated Molecular Mass ◽  
Reading Frame ◽  
E Coli ◽  
Type Iii

The gene catIII, encoding a type III enterobacterial chloramphenicol acetyltransferase, was cloned from the transmissible plasmid R387 into pBR322 and bacteriophage M13 mp8. Nucleotide sequence analysis of 1160 bp of DNA identified an open reading frame encoding a protein of 213 amino acid residues and a calculated molecular mass of 24965 Da. The predicted N-terminal sequence is identical with that determined by Edman degradation of chloramphenicol acetyltransferase purified from Escherichia coli harbouring R387. Sequences equivalent to the consensus motifs for initiation and rho-factor-independent termination of transcription in E. coli occur 5′ and 3′ to the catIII open reading frame. In contrast with the catI gene, present on transposon Tn9 and many enterobacterial plasmids, expression of catIII is not subject to cyclic AMP-mediated catabolite repression in vivo and there is no sequence in the 5′ non-coding DNA that resembles that deduced as the consensus for the binding of cyclic AMP receptor protein. Unique restriction-endonuclease cleavage sites were introduced adjacent to the catIII reading frame by using oligonucleotide-directed mutagenesis to facilitate insertion into E. coli expression vectors. Fully active chloramphenicol acetyltransferase represents 30-50% of the soluble protein component of cell-free extracts of E. coli containing the appropriate plasmids.

scholarly journals Cloning, Nucleotide Sequence, and Expression of the Gene Encoding the Bacteriocin Boticin B from Clostridium botulinum Strain 213B

2000 ◽  
Vol 66 (12) ◽  
pp. 5480-5483 ◽  
Author(s):  
Sean S. Dineen ◽  
Marite Bradshaw ◽  
Eric A. Johnson
Keyword(s):  
Amino Acids ◽  
Nucleotide Sequence ◽  
Clostridium Botulinum ◽  
Shuttle Vector ◽  
Open Reading Frame ◽  
Gene Encoding ◽  
Reading Frame ◽  
Heat Stable ◽  
Group I

ABSTRACT Boticin B is a heat-stable bacteriocin produced byClostridium botulinum strain 213B that has inhibitory activity against various strains of C. botulinum and related clostridia. The gene encoding the bacteriocin was localized to a 3.0-kb HindIII fragment of an 18.8-kb plasmid, cloned, and sequenced. DNA sequencing revealed the boticin B structural gene,btcB, to be an open reading frame encoding 50 amino acids. A C. botulinum strain 62A transconjugant containing theHindIII fragment inserted into a clostridial shuttle vector expressed boticin B, although at much lower levels than those observed in C. botulinum 213B. To our knowledge, this is the first demonstration and characterization of a bacteriocin from toxigenic group I C. botulinum.

scholarly journals Quasispecies of TT Virus (TTV) with Sequence Divergence in Hypervariable Regions of the Capsid Protein in Chronic TTV Infection

1999 ◽  
Vol 73 (11) ◽  
pp. 9604-9608 ◽  
Author(s):  
Tsutomu Nishizawa ◽  
Hiroaki Okamoto ◽  
Fumio Tsuda ◽  
Tatsuya Aikawa ◽  
Yoshiki Sugai ◽  
...  
Keyword(s):  
Amino Acids ◽  
Capsid Protein ◽  
Immune Surveillance ◽  
Sequence Divergence ◽  
Open Reading Frame ◽  
Amino Acid Substitutions ◽  
Reading Frame ◽  
Tt Virus ◽  
Hypervariable Regions ◽  
Open Reading Frame 1

ABSTRACT Three hypervariable regions were identified in a central portion of open reading frame 1 of TT virus DNA, which codes for a putative capsid protein of 770 amino acids. TT virus circulates as quasispecies, with many amino acid substitutions in hypervariable regions, to evade immune surveillance of the hosts and to establish a persistent infection.

DNA vaccination of pigs with open reading frame 1–7 of PRRS virus

Vaccine ◽  
2004 ◽  
Vol 22 (27-28) ◽  
pp. 3628-3641 ◽  
Author(s):  
Annette Malene Barfoed ◽  
Merete Blixenkrone-Møller ◽  
Merethe Holm Jensen ◽  
Anette Bøtner ◽  
Søren Kamstrup
Keyword(s):  
Dna Vaccination ◽  
Open Reading Frame ◽  
Reading Frame ◽  
Prrs Virus ◽  
Open Reading Frame 1
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